Cell fractionation and ultracentrifugation

Cell fractionation

Cells are broken open to release the content and organise are separated

cells are prepared in cold isotonic and buffered solution

Isotonic?

same water potential to prevent osmosis to prevent organism to shrivel or burst

Cold??

To reduce enzyme activity to prevent damage to organelles

Buffered??

Prevent damage to organelles

Cell fractionation steps

Step 1: Homogenisation- cells broken up using blender (cold isotonic buffered solution)

Step two: Filtration- filtered to remove debris

step 3: Ultracentrifugation-spins round at different speeds causing organelles to separate according to density

Ultracentrifugation

spun at low speed causing most dance organ to form bottom making a pellet the supernatant is removed and spun at a higher speed to form a pellet of the next most dense organells and so on

most dense Nucleus → chloroplast → mitochondria lysosomes→ endoplasmic reticulum → ribosomes least dense